Low-cost 3D-printed inverted microscope to detect Mycobacterium tuberculosis in a MODS culture.

MODS, an assay for diagnosis of tuberculosis and drug-susceptibility, is based in the microscopic observation of the characteristic cords of Mycobacterium tuberculosis colonies grown in liquid media. An inverted optical microscope (100× magnification) is required to observe and interpret MODS cultures. Unfortunately, the cost of commercial inverted microscopes is not affordable in low resource settings. To perform a diagnosis of tuberculosis using the MODS assay, images with modest quality are enough for proper interpretation. Therefore, the use of a high cost commercial inverted optical microscope is not indispensable. In this study, we designed a prototype of an optical inverted microscope created by 3D-printing and based on a smartphone. The system was evaluated with 226 MODS TB positive and 207 MODS TB negative digital images. These images were obtained from 10 sputum samples MODS positive and 10 sputum samples MODS negative. The quality of all images was assessed by a qualified technician, in terms of adequacy to interpret and classify them as positive or negative for tuberculosis. The quality of the images was considered appropriate for MODS interpretation. All the 20 samples were correctly classified (as TB positive/negative) by reading with the prototype 3D-printed inverted microscope.

Cough dynamics in adults receiving tuberculosis treatment.

Cough is a characteristic symptom of tuberculosis, is the main cause of transmission, and is used to assess treatment response. We aimed to identify the best measure of cough severity and characterize changes during initial tuberculosis therapy. We conducted a prospective cohort of recently diagnosed ambulatory adult patients with pulmonary tuberculosis in two tertiary hospitals in Lima, Peru. Pre-treatment and five times during the first two months of treatment, a vibrometer was used to capture 4-hour recordings of involuntary cough. A total of 358 recordings from 69 participants were analyzed using a computer algorithm. Total time spent coughing (seconds per hour) was a better predictor of microbiologic indicators of disease severity and treatment response than the frequency of cough episodes or cough power. Patients with prior tuberculosis tended to cough more than patients without prior tuberculosis, and patients with tuberculosis and diabetes coughed more than patients without diabetes co-morbidity. Cough characteristics were similar regardless of HIV co-infection and for drug-susceptible versus drug-resistant tuberculosis. Tuberculosis treatment response may be meaningfully assessed by objectively monitoring the time spent coughing. This measure demonstrated that cough was increased in patients with TB recurrence or co-morbid diabetes, but not because of drug resistance or HIV co-infection.

Food Choice and Dietary Intake among People with Tuberculosis in Peru: Implications for Improving Practice.

BACKGROUND: In Peru, tuberculosis (TB) is perceived as a nutritional disease. This perception, alongside factors including household food insecurity, may drive the food choices of people with TB and influence treatment outcomes. OBJECTIVES: The objective of this qualitative study was to explore drivers of food choice among adults recently diagnosed with TB. METHODS: The study was conducted between April and December 2016 in the Huaycán district of Lima, Peru. Structured questionnaires were administered to 39 adults with TB at the time of diagnosis and after 1 mo of treatment to characterize food security and socioeconomic status. At 1 mo of treatment, 24-h dietary recalls, enhanced by recipes obtained from local street vendors, were administered to examine patterns of food consumption and determine mean daily intake of macro- and micronutrients. Among a subset of 9 participants, in-depth interviews were used to explore dietary beliefs and food choices associated with TB. RESULTS: Overall, 13.2% of participants were underweight at baseline, and 10.5% were overweight. At 1 mo of treatment, the mean caloric intake was 600 kcal/d over what was needed to maintain their current weight. Most of these additional kilocalories came from carbohydrates. Patients made active efforts to improve their diets during treatment, and were both receptive to, and actively sought out, nutritional advice. However, many patients reported significant unnecessary spending on questionable commercial products, such as expensive natural remedies and nutritional supplements. CONCLUSIONS: The perceived connection between TB and diet creates both opportunities and challenges for treatment providers. Nutritional counseling provided through the national TB program should promote dietary quality through foods that are locally available, inexpensive, and aligned with cultural perceptions of health and wellness.

A novel inexpensive electrochemical sensor for pyrazinoic acid as a potential tool for the identification of pyrazinamide-resistant Mycobacterium tuberculosis.

Introduction: Tuberculosis (TB) is a significant cause of morbidity and mortality worldwide. The patient compliance with the long treatment regimens is essential for successful eradication. Pyrazinamide (PZA) shortens these regimens from 9 to 6 months, and therefore, improves treatment completion rates. Although PZA is a first-line medication for the treatment of TB, no simple or reliable assay to determine PZA resistance is yet available. In the presence of PZA, only susceptible Mycobacterium tuberculosis strains release pyrazinoic acid (POA). Therefore, the measurement and quantification of released POA is an indicator of PZA resistance. Methods: Two electrochemical sensors were constructed and tested with alternative working electrodes in conjunction with a portable potentiostat to measure the current produced when a potential difference of 2 V is applied to varying concentrations of POA in controlled solutions. Results: The large (13.2 mm) electrochemical sensor was able to detect POA at a minimum concentration of 40 µM to a statistically significant level (P = 0.0190). Similar graphical trends were obtained when testing the electrochemical sensor in the supernatant of a negative microscopic observation drug susceptibility assay culture, irrespective of the presence of PZA. Conclusion: Inexpensive and reusable electrochemical sensors with a portable potentiostat are a promising tool for the detection of POA, a biomarker of PZA susceptible M. Tuberculosis.

Cough Frequency During Treatment Associated With Baseline Cavitary Volume and Proximity to the Airway in Pulmonary TB.

BACKGROUND: Cough frequency, and its duration, is a biomarker that can be used in low-resource settings without the need of laboratory culture and has been associated with transmission and treatment response. Radiologic characteristics associated with increased cough frequency may be important in understanding transmission. The relationship between cough frequency and cavitary lung disease has not been studied. METHODS: We analyzed data in 41 adults who were HIV negative and had culture-confirmed, drug-susceptible pulmonary TB throughout treatment. Cough recordings were based on the Cayetano Cough Monitor, and sputum samples were evaluated using microscopic observation drug susceptibility broth culture; among culture-positive samples, bacillary burden was assessed by means of time to positivity. CT scans were analyzed by a US-board-certified radiologist and a computer-automated algorithm. The algorithm evaluated cavity volume and cavitary proximity to the airway. CT scans were obtained within 1 month of treatment initiation. We compared small cavities (≤ 7 mL) and large cavities (> 7 mL) and cavities located closer to (≤ 10 mm) and farther from (> 10 mm) the airway to cough frequency and cough cessation until treatment day 60. RESULTS: Cough frequency during treatment was twofold higher in participants with large cavity volumes (rate ratio [RR], 1.98; P = .01) and cavities located closer to the airway (RR, 2.44; P = .001). Comparably, cough ceased three times faster in participants with smaller cavities (adjusted hazard ratio [HR], 2.89; P = .06) and those farther from the airway (adjusted HR, 3.61;, P = .02). Similar results were found for bacillary burden and culture conversion during treatment. CONCLUSIONS: Cough frequency during treatment is greater and lasts longer in patients with larger cavities, especially those closer to the airway.

Dynamics of Cough Frequency in Adults Undergoing Treatment for Pulmonary Tuberculosis.

Background: Cough is the major determinant of tuberculosis transmission. Despite this, there is a paucity of information regarding characteristics of cough frequency throughout the day and in response to tuberculosis therapy. Here we evaluate the circadian cycle of cough, cough frequency risk factors, and the impact of appropriate treatment on cough and bacillary load. Methods: We prospectively evaluated human immunodeficiency virus-negative adults (n = 64) with a new diagnosis of culture-proven, drug-susceptible pulmonary tuberculosis immediately prior to treatment and repeatedly until treatment day 62. At each time point, participant cough was recorded (n = 670) and analyzed using the Cayetano Cough Monitor. Consecutive coughs at least 2 seconds apart were counted as separate cough episodes. Sputum samples (n = 426) were tested with microscopic-observation drug susceptibility broth culture, and in culture-positive samples (n = 252), the time to culture positivity was used to estimate bacillary load. Results: The highest cough frequency occurred from 1 pm to 2 pm, and the lowest from 1 am to 2 am (2.4 vs 1.1 cough episodes/hour, respectively). Cough frequency was higher among participants who had higher sputum bacillary load (P < .01). Pretreatment median cough episodes/hour was 2.3 (interquartile range [IQR], 1.2-4.1), which at 14 treatment days decreased to 0.48 (IQR, 0.0-1.4) and at the end of the study decreased to 0.18 (IQR, 0.0-0.59) (both reductions P < .001). By 14 treatment days, the probability of culture conversion was 29% (95% confidence interval, 19%-41%). Conclusions: Coughs were most frequent during daytime. Two weeks of appropriate treatment significantly reduced cough frequency and resulted in one-third of participants achieving culture conversion. Thus, treatment by 2 weeks considerably diminishes, but does not eliminate, the potential for airborne tuberculosis transmission.

Implementation y evaluación de una nariz electrónica para la detección de alcoholes lineales / Implementation and evaluation of an electronic nose for the alcohols detection / Implementação e avaliação de um nariz eletrônico para detectar álcoois lineares

Se desarrolló una nariz electrónica que permite la detección de alcoholes de manera sencilla y económica en comparación con las narices electrónicas tradicionales. Estábasada en cuatro sensores de gas de SnO2 (dos comerciales y dos fabricados en el laboratorio), un sistema neumático irregular, un hardware y software para adquisición de datos y un software de reconocimiento de patrones. Se evaluó el comportamiento de la nariz y las condiciones de trabajo con muestras de vapor de alcoholes (metanol, etanol, n-butanol y 1-2 octanol) y se determinó que los alcoholes se pueden detectar con el arreglo de sensores preparado y pueden diferenciarse entre sí haciendo uso del análisis estadístico de componentes principales (PCA). El orden de detección encontrado para los alcoholes lineales fue el siguiente: metanol > etanol > n-butanol > 1-octanol. Se encontró también que haciendo uso del análisis de componentes principales (PCA) y realizando una normalización de los datos en el software de reconocimiento de patrones, la varianza total de las muestras también aumenta del 76% al 85%. Esto demuestra que una nariz simple y económica puede clasificar bien las muestras evaluadas.

An electronic nose for the detection of alcohols, easy to use and inexpensive as compared to traditional electronic noses, was developed. This nose is based on four SnO2 gas sensors (two commercial and two homemade), an irregular pneumatic system, hardware and a software for data acquisition and a software for pattern recognition. The nose behavior and the working conditions with vapor samples of alcohols (methanol, ethanol, n-butanol and 1-octanol) were evaluated. Alcohols could be detected with the array of prepared sensors and could be also differentiated from each other by using principal component analysis (PCA). The detection order for linear alcohols followed the order: methanol > ethanol > n-butanol > 1-octanol. It was also found that by using PCA and performing a standardization of data in software pattern recognition so, the total variance of such information increases from 76% up to 85%. This result confirms that a simple and inexpensive nose can rank well the tested samples.

Foi feito um nariz eletrônico que permitiu a detecção de álcoois de jeito simplis e económico em comparacao com os narices electrónicos tradicionais. Estábaseado em quatro sensores de gás SnO2 (dois comerciais e dois feitos no laboratório), um sistema pneumático irregular, hardware e software para aquisição de dados e software de reconhecimento de padrões. Foi avaliado o comportamento do nariz e as condições de trabalho com amostras de vapor de álcoois (metanol, etanol, n-butanol e 1-octanol) determinou-se que os álcoois podem ser detectados com o arranjo de sensores preparado e pode diferenciar-se entre sim usando análise de componentes principais (PCA). A ordem de detecção para os álcoois lineares encontrados foi os seguinte: metanol> etanol> n-butanol> 1-octanol. Verificou-se que, usando análise de componentes principais (PCA) e a execução de uma normalização dos dados no software de reconhecimento de padrões, a variância das amostras também aumenta de 76% para 85%. Esto demostra que um nariz simples e barato pode classificar bem as amostras avaliadas.

Protocol for studying cough frequency in people with pulmonary tuberculosis.

INTRODUCTION: Cough is a key symptom of tuberculosis (TB) as well as the main cause of transmission. However, a recent literature review found that cough frequency (number of coughs per hour) in patients with TB has only been studied once, in 1969. The main aim of this study is to describe cough frequency patterns before and after the start of TB treatment and to determine baseline factors that affect cough frequency in these patients. Secondarily, we will evaluate the correlation between cough frequency and TB microbiological resolution. METHODS: This study will select participants with culture confirmed TB from 2 tertiary hospitals in Lima, Peru. We estimated that a sample size of 107 patients was sufficient to detect clinically significant changes in cough frequency. Participants will initially be evaluated through questionnaires, radiology, microscopic observation drug susceptibility broth TB-culture, auramine smear microscopy and cough recordings. This cohort will be followed for the initial 60 days of anti-TB treatment, and throughout the study several microbiological samples as well as 24 h recordings will be collected. We will describe the variability of cough episodes and determine its association with baseline laboratory parameters of pulmonary TB. In addition, we will analyse the reduction of cough frequency in predicting TB cure, adjusted for potential confounders. ETHICS AND DISSEMINATION: Ethical approval has been obtained from the ethics committees at each participating hospital in Lima, Peru, Asociación Benéfica PRISMA in Lima, Peru, the Universidad Peruana Cayetano Heredia in Lima, Peru and Johns Hopkins University in Baltimore, USA. We aim to publish and disseminate our findings in peer-reviewed journals. We also expect to create and maintain an online repository for TB cough sounds as well as the statistical analysis employed.

Towards autonomous lab-on-a-chip devices for cell phone biosensing.

Modern cell phones are a ubiquitous resource with a residual capacity to accommodate chemical sensing and biosensing capabilities. From the different approaches explored to capitalize on such resource, the use of autonomous disposable lab-on-a-chip (LOC) devices-conceived as only accessories to complement cell phones-underscores the possibility to entirely retain cell phones' ubiquity for distributed biosensing. The technology and principles exploited for autonomous LOC devices are here selected and reviewed focusing on their potential to serve cell phone readout configurations. Together with this requirement, the central aspects of cell phones' resources that determine their potential for analytical detection are examined. The conversion of these LOC concepts into universal architectures that are readable on unaccessorized phones is discussed within this context.

Empleo de algoritmos matemáticos para la evaluación de la influencia de los parámetros fisicoquímicos que afectan la adsorción de compuestos aromáticos sobre carbón activado / Use of mathematical algorithms to evaluate the influence of physicochemical parameters affecting the adsorption of aromatic compounds on activated carbon / Uso de algoritmos matemáticos para avaliar a influência dos parâmetros físico-químicos que afetam a adsorção de compostos aromáticos sobre carbono ativado

El objetivo principal fue encontrar cómo ciertos parámetros o factores fisicoquímicos del carbón activado pueden influir en la capacidad de adsorción de tres adsorbatos: fenol, ácido benzoico y ácido salicílico. Se emplearon dos métodos de análisis multivariado de datos: análisis principal de mínimos cuadrados (PLS) y regresión de componentes principales (PCR). El método de PLS mostró una mejor concordancia entre los valores estimados y experimentales. Usando este método, se formularon ecuaciones para predecir la capacidad de remoción de cada adsorbato. Usando PLS fue posible estimar la capacidad de adsorción del ácido benzoico, ácido salicílico y fenol con un error estándar de validación menor al 6%. Así se predijo que la acidez superficial es el parámetro más importante del carbón activado para adsorber compuestos aromáticos.

The main objective was to describe parameters and physicochemical factors of activated carbon related to the adsorption capacity of three adsorbates: Phenol, benzoic acid, and salicylic acid. Two multivariate data analysis methods were used: Partial least square (PLS) and principal component regression (PCR). PLS showed better agreement between estimated and experimental values and using this method, equations were developed to predict the removal capacity of each adsorbate. The adsorption capacity of activated carbon in relation to benzoic acid, salicylic acid, and phenol was predicted with a standard error of validation of less than 6%. Surface acidity was the most important parameter affecting the adsorption of aromatic compounds by activated carbon.

Foram utilizados métodos de análises multivariada de dados: análises parcial de mínimos quadrados (PLS) e regressão das componentes principais (PCR) para encontrar como certos parâmetros ou fatores físico-químicos do carvão ativado podem influenciar a capacidade de adsorção de três adsorvatos: fenol, ácido benzoico e ácido salicílico. Encontrou-se que o método PLS apresentou melhor concordância entre os valores estimados e experimentais. Utilizando-se este método, podem-se desenvolver equações para prever a capacidade de remoção para cada adsorvato. Usando o método PLS foi possível estimar a capacidade de adsorção do carvão ativado para o ácido benzoico, ácido salicílico e fenol com um erro padrão de validação menor ao 6%. Assim foi previsto que a acidez da superfície é o parâmetro mais importante do carvão ativado para adsorver compostos aromáticos.

Autonomous Chemical Sensing Interface for Universal Cell Phone Readout.

Exploiting the ubiquity of cell phones for quantitative chemical sensing imposes strong demands on interfacing devices. They should be autonomous, disposable, and integrate all necessary calibration and actuation elements. In addition, a single design should couple universally to a variety of cell phones, and operate in their default configuration. Here, we demonstrate such a concept and its implementation as a quantitative glucose meter that integrates finger pumps, unidirectional valves, calibration references, and focusing optics on a disposable device configured for universal video acquisition.

Low cost lab-on-a-chip prototyping with a consumer grade 3D printer.

Versatile prototyping of 3D printed lab-on-a-chip devices, supporting different forms of sample delivery, transport, functionalization and readout, is demonstrated with a consumer grade printer, which centralizes all critical fabrication tasks. Devices cost 0.57US$ and are demonstrated in chemical sensing and micromixing examples, which exploit established principles from reference technologies.

PDMS lab-on-a-chip fabrication using 3D printed templates.

The fabrication of conventional PDMS on glass lab-on-a-chip (LOC) devices, using templates printed with a commercial (2299 US$) micro-stereo lithography 3D printer, is demonstrated. Printed templates replace clean room and photolithographic fabrication resources and deliver resolutions of 50 µm, and up to 10 µm in localized hindrances, whereas the templates are smooth enough to allow direct transfer and proper sealing to glass substrates. 3D printed templates accommodate multiple thicknesses, from 50 µm up to several mm within the same template, with no additional processing cost or effort. This capability is exploited to integrate silicone tubing easily, to improve micromixer performance and to produce multilevel fluidics with simple access to independent functional surfaces, which is illustrated by time-resolved glucose detection. The templates are reusable, can be fabricated in under 20 min, with an average cost of 0.48 US$, which promotes broader access to established LOC configurations with minimal fabrication requirements, relieves LOC fabrication from design skills and provides a versatile LOC development platform.

Validation of an automated cough detection algorithm for tracking recovery of pulmonary tuberculosis patients.

BACKGROUND: A laboratory-free test for assessing recovery from pulmonary tuberculosis (TB) would be extremely beneficial in regions of the world where laboratory facilities are lacking. Our hypothesis is that analysis of cough sound recordings may provide such a test. In the current paper, we present validation of a cough analysis tool. METHODOLOGY/PRINCIPAL FINDINGS: Cough data was collected from a cohort of TB patients in Lima, Peru and 25.5 hours of recordings were manually annotated by clinical staff. Analysis software was developed and validated by comparison to manual scoring. Because many patients cough in bursts, coughing was characterized in terms of cough epochs. Our software correctly detects 75.5% of cough episodes with a specificity of 99.6% (comparable to past results using the same definition) and a median false positive rate of 4 false positives/hour, due to the noisy, real-world nature of our dataset. We then manually review detected coughs to eliminate false positives, in effect using the algorithm as a pre-screening tool that reduces reviewing time to roughly 5% of the recording length. This cough analysis approach provides a foundation to support larger-scale studies of coughing rates over time for TB patients undergoing treatment.

Cough detection algorithm for monitoring patient recovery from pulmonary tuberculosis.

In regions of the world where tuberculosis (TB) poses the greatest disease burden, the lack of access to skilled laboratories is a significant problem. A lab-free method for assessing patient recovery during treatment would be of great benefit, particularly for identifying patients who may have drug-resistant tuberculosis. We hypothesize that cough analysis may provide such a test. In this paper we describe algorithm development in support of a pilot study of TB patient coughing. We describe several approaches to event detection and classification, and show preliminary data which suggest that cough count decreases after the start of treatment in drug-responsive patients. Our eventual goal is development of a low-cost ambulatory cough analysis system that will help identify patients with drug-resistant tuberculosis.

Development of low-cost inverted microscope to detect early growth of Mycobacterium tuberculosis in MODS culture.

BACKGROUND: The microscopic observation drug susceptibility (MODS) assay for rapid, low-cost detection of tuberculosis and multidrug resistant tuberculosis depends upon visualization of the characteristic cording colonies of Mycobacterium tuberculosis in liquid media. This has conventionally required an inverted light microscope in order to inspect the MODS culture plates from below. Few tuberculosis laboratories have this item and the capital cost of $5,000 for a high-end microscope could be a significant obstacle to MODS roll-out. METHODOLOGY: We hypothesized that the precise definition provided by costly high-specification inverted light microscopes might not be necessary for pattern recognition. SIGNIFICANCE: In this work we describe the development of a low-cost artesenal inverted microscope that can operate in both a standard or digital mode to effectively replace the expensive commercial inverted light microscope, and an integrated system that could permit a local and remote diagnosis of tuberculosis.